Opaque additive to block stray light in teflon® af light-guiding flowcells

ABSTRACT

The present invention is directed to the use of a light absorbing wall material to eliminate stray light paths in light-guiding structures, such as those used for HPLC absorbance detection. More specifically, the present invention relates to the use of carbon-doped Teflon® AF, or “black Teflon® AF,” for all or part of the walls of a light-guiding flowcell adapted for use in HPLC absorbance detection.

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims priority from a PCT International PatentApplication No. PCT/US03/05811, filed Feb. 25, 2003 and U.S. ProvisionalPatent Application No. 60/359,354, filed Feb. 25, 2002. The contents ofthese applications are incorporated herein by reference.

BACKGROUND OF THE INVENTION

The present invention relates generally to the use of a light absorbingwall material to eliminate stray light paths in light-guidingapplications, such as High Performance Liquid Chromatography (HPLC), andCapillary Zone Electrophoresis (CZE) spectroscopic analysis.

Systems for light absorption detection generally comprise four basiccomponents; a light source, a means for selecting wavelengths to beused, a light-guiding vessel, typically in the form of a hollow tube orcapillary through which a sample to be analyzed and light are passed (aflowcell), and a light detector which measures the amount of lighttransmitted through the flowcell. Large optical throughput can beachieved when the light is guided along the capillary similar to the waylight is guided along an optical fiber.

A flowcell must be constructed from materials that are resistant to thesolutions encountered in liquid chromatography or CE. To achieve highsensitivity to small concentrations of analyte, the cell must have ahigh optical throughput and a long pathlength. If the quantity ofanalyte is small and capillary separation techniques are used, thevolume of the cell must also be small, otherwise band spreading and lossof chromatographic resolution occurs. The transmittance, T, of lightthrough such a system filled with a light absorbing sample is determinedin accordance with Beer's law: $\begin{matrix}{T = \frac{I}{I_{0}}} & \left( {1a} \right) \\{A = {{\log_{10}\left( \frac{I_{0}}{I} \right)} = {ɛ\quad{bc}}}} & \left( {1b} \right)\end{matrix}$

-   -   where I₀ is the light exiting the flowcell when it is filled        with clear mobile phase and I is the light power exiting the        flowcell when analyte is present. b is the path length of the        flowcell conventionally expressed in centimeters, c is the        analyte concentration in M or moles/liter and ε is the molar        absorptivity expressed in units of cm⁻¹(moles/liter)⁻¹. A is the        absorbance, a dimensionless number expressed in absorbance units        (au).

The requirement for high light throughput and long pathlength isillustrated by differentiating equation (1b). $\begin{matrix}{{\Delta\quad c} = \frac{\Delta\quad A}{b\quad ɛ}} & \left( {1c} \right)\end{matrix}$

Δc represents the smallest analyte concentration that can be detectedand ΔA the corresponding smallest change in absorbance that can bemeasured. This represents the noise at the absorbance baseline, theoutput of the absorbance detector.

As illustrated by equation (1b), low absorbance noise requires a highlight signal 10 and low noise in the measurement of I, i.e. a highsignal-to-noise (S/N) ratio in the raw transmittance measurement. In awell-designed detector, shot noise, which is proportional to the squareroot of the light signal, dominates, so high SIN requires high lightthroughput.

Light-guiding flowcells enable low volume cells to be constructed withhigh light throughput and long path length. The liquid sample iscontained in a tube of material having a lower refractive index (RI)than the mobile phase. Light is introduced into one end of the tube andpropagates down the axis of the tube making multiple internalreflections before emerging at the other end. The liquid is analogous tothe core of an optical fiber and the material of the tube is analogousto the cladding. The condition for light guiding is that the raysincident on the liquid/wall boundary do so at an angle of incidencegreater than the critical angle θ_(c). $\begin{matrix}{\theta_{c} = {\sin^{- 1}\frac{n_{2}}{n_{1}}}} & \left( {2a} \right)\end{matrix}$where n₁ is the RI of the liquid, and n₂ is the RI of the wall of theflowcell.

The numerical aperture (NA) of the guided beam is given by:NA=sin⁻¹φ=(n ₁ ² −n ₂ ²)^(1/2)  (2b)Where φ is the largest angle, between a ray entering the cell from airand the cell axis, which meets the guiding condition. The guidingmechanism is termed total internal reflection (TIR)

Recently, flowcells having an inner surface of an amorphousfluoropolymer material that has an index of refraction lower than thatof common chromatography solvents, e.g. water, have enabledlight-guiding flowcells to be constructed.

Light introduced along the axis of the tube is guided in the fluid bytotal internal reflection at the fluid wall boundary. The amorphousfluoropolymer material Teflon® AF 1600 and 2400 are preferred tubematerials because they are transparent throughout the visible andultraviolet spectrum, they have an unusually low refractive index (1.31and 1.29 respectively) and are chemically inert. As a comparison, the RIof water at the same wavelength is 1.333. All common solvents (asmethanol/water mixtures and acetonitrile) have a higher RI than waterand therefore also Teflon® AF. Only pure methanol has an index slightlybelow water, but still above that of Teflon® AF. Even at differentwavelengths, the fluoropolymers retain the RI advantage.

However, it is difficult to construct a cell with amorphousfluoropolymer walls without some light entering the end cross-section ofthe wall, or some light 18 being scattered into the wall from theliquid, or some light 18 entering the fluid from the walls afterbypassing part or all of the sample fluid. These aberrant light pathsresult in a stray light background and inaccuracy in the readings,limiting the linearity and dynamic range of a detector that is supposedto receive only light that has passed through the liquid.

One strategy to control stray light positions opaque masks between thewalls and the light, but the small diameter tubes of HPLC and CZEequipment makes the alignment of such masks difficult and timeconsuming. A second strategy to control stray light supplies enteringlight through an optical fiber having an OD that fits between the walls,but the amount of light coupled into the liquid is reduced geometricallyby the reduced area

The difficulty of controlling stray light becomes greater as fluidcross-sections are made smaller. For capillary HPLC or CZE detection, afluid channel ID of 100 μm or less is needed to create a small volumeflowcell, with sufficient pathlength to preserve analytical sensitivity.A better way is needed to fabricate light guiding flowcells to avoid thedifficulties of controlling stray light outlined above

SUMMARY OF THE INVENTION

The present invention provides an apparatus and method for controllingstray light in vessels having small cross sections, used especially forlight absorption measurements.

In a preferred application, the vessel is a flowcell receiving analytefrom a HPLC apparatus. One embodiment is a device for receiving one ormore samples and measuring light emitted or refracted therefrom. Thisdevice comprises a vessel with a cavity for containing a sample during ameasuring process, where the vessel comprises at least one wall in fluidcontact with the sample, the wall having a composition having an indexof refraction lower than the index of refraction of a fluid in thesample. Further the wall has an absorption coefficient sufficient tosubstantially attenuate light propagating through the wall and thevessel has at least one means for passage of transmitting light, whereinlight within the cavity is guided by attenuated total reflection intothe sample. By judicious choice of the wall absorption coefficient, thelight guided through the fluid by internal reflection is only minimallyattenuated. The means for means for passage of transmitting light may bean opening, a window, a lens or an optical fiber. A preferred materialfor the wall is Teflon® AF fluoropolymer doped with a black dopant suchas carbon black.

Another embodiment is a light-guiding apparatus comprising a fluidchannel bounded by at least one wall, the composition of the wall havingan index of refraction lower than the index of refraction of a fluid inthe fluid channel and having an absorption coefficient sufficient tosubstantially attenuate light propagating through the walls. Theapparatus further comprises an entrance and exit for light, the entranceand exit perpendicular to an axis of the channel and fluid inlet andoutlet ports, whereby the walls and fluid effect guiding of the light byattenuated total reflection, the walls having minimal absorptive effecton the guided light. The walls have an absorption coefficient in therange of 0.1 to 100 mm⁻¹ at a wavelength within the wavelength range ofUV, visible and near IR. A preferred material for the walls is Teflon®AF fluoropolymer doped with a black dopant such as carbon black. Aconcentration of carbon black between 0.01% and 1% by weight of thefluoropolymer is sufficient to absorb stray light.

Another embodiment is an apparatus for housing a liquid sample and forexposing the liquid sample to light. The apparatus comprises a conduithaving a wall formed of an amorphous fluoropolymer having a refractiveindex less than the refractive index of water and having an absorptioncoefficient of a magnitude such that when the conduit is filled withwater, visible and ultra-violet light can be transmitted, substantiallywithout loss, along the axis of the conduit by attenuated totalreflection but visible and ultra-violet light are substantiallycompletely absorbed in passage through the walls of the conduit.

A method of performing photometric analysis of a liquid sample withimproved linearity of detection comprises introducing the liquid sampleinto a conduit having a wall formed of an amorphous fluoropolymer havinga refractive index less than a refractive index of water and having anabsorption coefficient sufficient to substantially attenuate lightpropagation through the wall, shining light axially onto the conduitfilled with sample liquid, receiving light transferred through theliquid sample at a detector; and determining the concentration of thesample in the liquid by measuring the light absorption of the sample.When light is axially shone onto the conduit filled with sample liquid,light transferred through the liquid sample is detected and theconcentration of sample in the liquid is determined. Alternately thesample concentration can be determined used the emitted florescence orRaman scattered light.

A set of light-guiding flowcells for a fluid comprises a set of channelsformed in a substrate of material having an index of refraction lowerthan the index of the fluid and an absorption coefficient sufficient tosubstantially attenuate light propagating through the material. A set ofchannel covers formed in a section of material having an index ofrefraction and absorption coefficient identical to the substrate ofmaterial is fixed to the set of channels forming a set of coveredchannels. At least one covered channel, or interconnected set of coveredchannels, has a fluid inlet and outlet port. At least one coveredchannel has a source of light at a light entrance end and has a lightexit end. The at least one covered channel and fluid therefore effectguiding of the light by attenuated total reflection and the at least onecovered channel has minimal absorptive effect on the light guided byinternal reflection. A covered channel may be configured as a separationcolumn. The interconnected channels for perform analysis on a fluidpassing therethrough when the output of the separation column isconnected to the light-guiding channel, where the light exit end isconnected to a detector external to the set of flowcells.

A typical use for the invention is in a flowcell for HPLC or CEabsorbance detection where the flowcell is constructed from a hollowtube of low index material such as Teflon® AF 2400, darkened wherein thewall material is sufficiently absorbing to block light transmissionthrough the wall. At the same time, the absorption by the wall is lowenough that light guiding is substantially unaffected. Carbon-dopedTeflon® AF, or “black Teflon® AF,” is a material well adapted for all orpart of the walls of such a light-guiding flowcell for use in HPLCabsorbance detection

BRIEF DESCRIPTION OF THE DRAWINGS

These and other features and advantages of the present invention willbecome more apparent from the following detailed description taken inconjunction with the accompanying drawings wherein like referencecharacters denote corresponding parts throughout the several views, andwherein:

FIG. 1. is a simplified diagram of a light guiding vessel according tothe invention;

FIG. 2. is a simplified diagram of the light-guiding vessel in the formof a flowcell according to the invention;

FIG. 3. is a graph illustrating the depth of penetration of a light waveinto a wall according to the invention;

FIG. 4. is a diagram of a flowcell constructed incorporating theinvention; and

FIG. 5 is a simplified diagram of a “lab-on-a chip” implementedutilizing the invention.

DETAILED DESCRIPTION

A light-guiding structure that has superior signal to noisecharacteristics can be constructed if the absorption coefficient of thewalls of the liquid containing vessel are judiciously chosen to onlyminimally attenuate the light guided through the liquid by internalreflection. The vessel may be formed to contain a single sample, as anarray of single sample vessels or as flowcell capable of analyzing asequence of samples. The basis of the invention is discussed utilizingas an example a light guiding flowcell which is simple and easy toconstruct, minimizes stray light and exhibits maximum light throughputfor a chosen volume and pathlength. The low RI wall material is madesufficiently absorbing to block stray light paths that bypass part of orall of the liquid sample, a cause of nonlinearity between absorbance andconcentration. The absence of masks or optical fibers inserted into theflowcell lumen allows optical throughput to be maximized.

The following discussion justifies the assertion that a bulk absorptioncoefficient for the low index wall material can be chosen such thatundesired rays that enter the wall are absorbed, and yet desired rays,which are guided along the flowcell channel, are transmitted withoutloss. Absorption of light through a material is described by theBouguer/Lambert law: $\begin{matrix}{\frac{I}{I_{0}} = {\mathbb{e}}^{- {ab}}} & (3)\end{matrix}$where α is the material absorption coefficient in units of reciprocallength, b is the distance traveled in the material and, ignoring endreflection losses, I₀ and I are the light intensity, or power, enteringand leaving the material respectively.

Adequate light blocking can be achieved if the light transmitted throughthe tube wall, parallel to the lumen, is reduced to one part in 1,000 ofthe incident intensity, within a distance of 5 mm. This essentiallyeliminates stray light through the bulk material. With these parameters,Equation (3) yields an absorption coefficient of α=1.4 mm⁻¹. The effectof this change in the material on the light-guiding properties isconsidered below.

When light rays are guided by total internal reflection at the boundarybetween core material and a lower index cladding, there is a smallpenetration of light into the low index medium. If the low index mediumis transparent, the internal reflection is 100%. But, if the low indexmedium absorbs light, some energy is trapped in the penetratingevanescent light wave and the process is referred to as attenuated totalreflection (ATR). When absorption is low, as in the present design,relatively simple expressions can be used to calculate the effectivethickness of Teflon® AF, penetrated by the light at each internalreflection (see Harrick N.J., Internal Reflection Spectroscopy, HarrickScientific Corp., Ossining, N.Y., 1987, p 43). The effective thicknessdepends on the wavelength, angle of incidence, refractive index of thetwo media, and on the plane of polarization. The physical orientationsof the components are illustrated in FIG. 4.

For light polarized perpendicular to the plane of incidence effectivethickness is: $\begin{matrix}{d_{e{({perp})}} = \frac{\lambda_{1}n_{21}\cos\quad\theta}{\left( {{\pi\left( {1 - n_{21}^{2}} \right)}\left( {{\sin^{2}\theta} - n_{21}^{2}} \right)^{1/2}} \right)}} & \left( {4a} \right)\end{matrix}$

For light polarized parallel to the plane of incidence effectivethickness is: $\begin{matrix}{d_{e{({parallel})}} = \frac{\lambda_{1}n_{21}\cos\quad{\theta\left( {{2\sin^{2}\theta} - n_{21}^{2}} \right)}}{\left( {{\pi\left( {1 - n_{21}^{2}} \right)}\left\{ {{\left( {1 + n_{21}^{2}} \right)\sin^{2}\theta} - n_{21}^{2}} \right\}\left( {{\sin^{2}\theta} - n_{21}^{2}} \right)^{1/2}} \right)}} & \left( {4b} \right)\end{matrix}$where: n₁ is the refractive index of medium 1, the fluid sample

-   -   n₂ is the refractive index of medium 2, the Teflon® AF in        contact with the fluid θ is the angle of incidence of a ray        internally reflected at the boundary between the two    -   n₂₁=n₂/n₁    -   λ₁=λ/n₁ is the wavelength in medium 1. λ is the wavelength in        air.

One possible application utilizes a 5 mm long light-guiding flowcellhaving an ID of 100 μm, a numerical aperture (NA) of the tube of 0.27and a beam of light having a wavelength of 250 nm. The rays that makethe largest angle with the axis (arcsin(NA)), and reflect from theboundary closest to the critical angle, make about ten reflections andpenetrate the deepest into the wall. The effective thickness of Teflon®AF traversed by these rays is, assuming unpolarized light, the averageof equations (4a) and (4b) above. For the worst case ray, the effectivethickness is 1.4 μm per reflection or 14 μm for the full length of theflowcell. Using this value for b, and the value of a calculated above,equation (3) gives the transmittance of the guided light as 0.98. Raysmaking a smaller angle with the axis have an even higher transmittance.These calculations indicate that with this absorption coefficient, whichis sufficiently large to block light transmitted through the darkenedTeflon® AF wall, attenuation of the guided beam is negligible.

FIG. 1 illustrates a single sample vessel 2 constructed using theinvention for measuring light after it traverses the sample. The sampleto be tested is placed in the cavity 6 of the vessel 2. An means forpassage of light 10 is provided in the walls 4 of vessel 2 to allowlight to enter 16 and exit 16′. The means for passage of light may be anopening, a window, a lens or an optic fiber. Since the walls 4 areformed of darkened Teflon® AF, light having an angle of incidence lessthan the critical angle, is guided down the cavity and some of the lightis reflected off the end of the cavity to be measured as it exits thecavity. Light that impinges on the end surfaces 11 of the walls 4 isabsorbed and does not interfere with the measurement. This vessel can beadapted for other configurations. An array of vessels 2 can beconstructed to test multiple samples. If a window replaces reflectingsurface 8, the measurement sensor can be placed opposite the lightsource. This adaptation may prove advantageous for high volume screeningof discrete samples. When the vessel 2 is further modified to allowthrough passage of a sample fluid, a flowcell such as illustrated inFIG. 2 is formed.

FIG. 2 illustrates the simplified design of a flowcell made possibleusing this invention. Tube 17 has darkened Teflon® AF walls. Fluid undertest enters at one end 18 of tube 17 and exits at the other end 19.Incident light 16, 24 is oriented at an angle within the NA of tube 17.The source of light 19 is wider than the ID of the tube 17 and isaligned with the axis. Light 16 that is oriented to enter the liquidmedium having an RI of n₁ is guided down the cavity 15 because the angleof incidence, 0, is less than the critical angle. Light 24 that fallsoutside the cavity enters the cell walls 4, travels a small distance,and is absorbed. FIG. 2 shows a simple window 26 at the exit of thelight-guiding flowcell. This avoids matching and aligning an opticalfiber to the flowcell exit and leads to better optical efficiency. Theentrance to the flowcell can also be a window, with the light focussedon the entrance. Rays hitting the walls 4 if the tube 17 are absorbed asabove. The design is simplified because masks are no longer needed atthe end of the cell. Optical efficiency is maximized because the wholeof the fluid path cross-section can be flooded with light.

FIG. 3 shows the number of reflections (diamonds) made by a light waveundergoing total internal reflection from the inside wall of a Teflon®AF tube filled with water in the case calculated above to determine theeffect on transmittance. Also plotted is the “effective thickness”(squares) of Teflon® AF material traversed by the light during itspassage through the light guide. This enables the attenuation of a beamof light to be calculated as it makes multiple reflections down thetube. The data in FIG. 3 are plotted against numerical aperture, NA. NAdefines the cone of rays entering the light guide. The most inclined raymakes an angle sin⁻¹(NA) with the axis in air external to the lightguide. This is the guided ray which experiences the largest attenuationfrom wall absorption and is the one whose data are plotted in FIG. 3.

If ten times the concentration of black dopant were used, giving a wallabsorption coefficient of α=14 mm⁻¹, transmittance of the worst case raywould drop to 82%, a significant but tolerable loss. This allowsreasonable latitude in selecting the concentration of the opaque dopant,or alternatively, allows the dopant absorption coefficient to vary by afactor of ten over the desired wavelength range and still meet thenecessary criteria.

A preferred opaque dopant is chemically resistant to HPLC solvents andpH range, and has as flat a spectrum as possible over the wavelengthregion of interest, 200 to 800 nm. Carbon black has been incorporatedinto fused silica to make “black quartz” used to block stray light pathsin flowcells and cuvettes, particularly those used to analyzefluorescence (See Hulme, U.S. Pat. No. 5,493,405; Fujita et al., U.S.Pat. No. 6,106,777). In these cells, no light guiding is involvedbecause the wall material has a higher index than the analytical fluid.However, as a material which blocks transmitted light over a wide rangeof wavelengths, carbon is a good choice for the present application tolight guided cells. Other materials could be used, such as finelydivided metal particles. Many different dopants could be used over morerestricted wavelength ranges.

FIG. 4 illustrates how a flowcell incorporating the invention may beconstructed. The flowcell body 42 is constructed from a sealablematerial such as PEEK in multiple parts. The tube of darkened Teflon® AF44 is supported in part of the cell body 42. Fluid 38 enters the bodyvia capillary 40 and is brought to the tube cavity 43 by channels 45etched in metal gaskets 32 between the body parts 42. After passingthrough the tube 44, fluid is further directed through the metal gaskets32 and out a fluid port 34. Light 36 enters the flowcell via an opticfiber 35 so as to traverse the fluid-filled tube cavity 43. At the exitside of the tube 44, a window 30 allows light 46 to exit the flowcelltoward the sensor. It is evident that light passes along the length ofthe light guiding tube 44. In this flowcell, the parts of the metalgaskets 32 that cover the ends of the tube walls 44 are not critical forlight blocking because the opacity of the walls of tube 44 preventsstray light from entering the fluid path.

In one embodiment, finely-divided carbon black is mixed with Teflon® AF2400 resin in powder form. The carbon black concentration is on theorder of 0.01%-1%. In a preferred embodiment the carbon blackconcentration is approximately 0.1%. The resultant mixture is used tomake “black” Teflon® AF tubing by extrusion or drawing usingwell-established methods. The low level of carbon black does notmaterially affect the ability to fabricate tubing. When the “black”Teflon® AF tubing is used in a light-guiding flowcell, it blocks thetransmission of light through the cell walls so none of that stray lightcan reach the detector and cause errors in the measurement.

“Lab-on-a-chip” structures, where certain channels are fluidicconnections, separation columns and reaction chambers have been producedon planar substrates. As a second embodiment of the present invention,strips of “black” Teflon® AF are extruded in ribbon form, several mmwide by about 1 mm thick. Sections of this are used as the substrate toform patterns of channels by, for example, hot embossing. Two suchsubstrates bonded together, or one with an unstructured lid, are used tocreate “lab-on-a-chip” structures with unique properties. Some channelsin the chip are used as fluidic connections to other channels formed asseparation columns or the like. These can be connected to fluidcontaining light-guides created for detection. Windows intolight-guiding sections are created and lengths of optical fiber bondedinto the sandwich to bring the needed light onto the chip for attachmentto the appropriate channel. Using opaque “black” Teflon® AF material asthe substrate blocks stray light and allows light-guiding detectionchannels to be constructed. In addition, the “black” Teflon® AF bulkmaterial prevents any stray light from the other sections of the chipfrom leaking light to the detection system. This improves detectorlinearity, as previously discussed. Use of the light-blocking substrateallows multiple light-guiding flowcells to be constructed on the samechip, without cross-talk.

FIG. 5 illustrates a simplified representation of a “lab-on-a-chip” madepossible by the invention. Substrate 50 is formed from “black” Teflon®AF. Channel 51 forms a fluidic connection from an entrance 52 to achannel 54 formed as a separation column. Light fibers 56 and 58 arebonded so as to form part of the end walls of light guiding channel 60used to measure the absorbance of a sample passing through channel 60.The fluid exits the structure through channel 62. On the same substrate,a separate light-guiding flowcell allows measurement the absorbance of afluid passing through channel 53 to channel 67 which is the measurementchannel. Separate light fibers 68, 70 are bonded to channel 67 to formthe light entrance and exit. Because of the opacity of substrate 50,light is confined to the two light-guiding measurement channels 60, 67.

While the application of light-blocking low RI material has beendiscussed in the context of absorption measurement, the material is alsoapplicable to light guiding applications used for fluorescence and Ramandetection. The opaque cell walls absorb unwanted excitation light whichenters or is scattered into the cell walls.

The material has been discussed in the context of a cuvette formed of anoptically transparent material drawn into the form of a cylindricalthin-walled capillary as used in absorbance measurement. However,further applications of the “black” Teflon® AF form the cuvette into acontainer with a reflective surface opposite a light enteringwindow/opening or a vessel with transparent windows opposite each other.

It is understood that various modifications may be made to theembodiments disclosed herein. Therefore, the above descriptions shouldnot be considered limiting, but merely as exemplifications of thevarious embodiments. Those skilled in the art will envision othermodifications within the scope and spirit of the claims appended hereto.

1. A device for receiving one or more samples and measuring lightemitted or refracted therefrom comprising: a vessel with a cavity forcontaining a sample during a measuring process, said vessel comprising:at least one wall in fluid contact with said sample, said wall having acomposition having an index of refraction lower than an index ofrefraction of a fluid in said sample and having an absorptioncoefficient sufficient to substantially attenuate light propagatingthrough said at least one wall; and at least one means for passage oftransmitting light; wherein light within said cavity is guided byattenuated total reflection into said sample.
 2. The device of claim 1wherein said vessel has at least one fluid inlet for receiving saidsample.
 3. The device of claim 1 wherein said vessel has at least onefluid outlet for discharging said sample.
 4. The device of claim 1wherein said means for passage of transmitted light is selected from thegroup comprising an opening, a window, a lens and an optic fiber.
 5. Thedevice of claim 1 further comprising a reflective surface opposite saidmeans for passage of transmitted light for returning said light that haspassed through said sample back to said means through said sample. 6.The device of claim 1 further comprising means for transmitting light tosaid sample and a window, opposite said means, for receiving light thathas passed through said sample.
 7. The device of claim 1 wherein alength of said cavity is between 10 and 50 times an inner diameter ofsaid cavity.
 8. The device of claim 1 wherein said walls are formed ofTeflon® AF fluoropolymer.
 9. The device of claim 8 wherein said Teflon®AF fluoropolymer is doped with black dopant.
 10. The device of claim 9wherein said black dopant is carbon black or finely divided metalparticles.
 11. The device of claim 8 wherein a concentration of saiddopant is between 0.01 and 0.1% of said fluoropolymer.
 12. The device ofclaim 8 wherein a concentration of said dopant is approximately 0.1% ofthe fluoropolymer.
 13. A light-guiding apparatus comprising: a fluidchannel bounded by at least one wall, the composition of said at leastone wall having an index of refraction lower than an index of refractionof a fluid in said fluid channel and having an absorption coefficientsufficient to substantially attenuate light propagating through saidwalls; an entrance for light, said entrance perpendicular to an axis ofsaid channel; an exit for light, said exit perpendicular to said axis;and fluid inlet and outlet ports, whereby said walls and fluid effectguiding of said light by attenuated total reflection, said walls havingminimal absorptive effect on the guided light.
 14. The light-guidingapparatus of claim 13 wherein said walls are formed in a hollow tube.15. The light-guiding apparatus of claim 13 wherein said walls have anabsorption coefficient in the range of 0.1 to 100 mm⁻¹ at a wavelengthwithin a wavelength range of UV, visible and near IR.
 16. Thelight-guiding apparatus of claim 13 wherein said walls are formed ofTeflon® AF fluoropolymer.
 17. The light-guiding apparatus of claim 16wherein said Teflon® AF is doped with black dopant.
 18. Thelight-guiding apparatus of claim 17 wherein said black dopant is carbonblack or finely divided metal particles.
 19. The light-guiding apparatusof claim 18 wherein a concentration of said black dopant is between0.01%-1% by weight of the fluoropolymer.
 20. The light-guiding apparatusof claim 19 wherein said dopant concentration is approximately 0.1% ofthe fluoropolymer.
 21. The light-guiding apparatus of claim 13 whereinthe apparatus is a flowcell.
 22. The light-guiding apparatus of claim 13wherein said fluid inlet and outlet ports are configured off said axis.23. An apparatus for housing a liquid sample and for exposing saidliquid sample to light, said apparatus comprising: a conduit having awall formed of an amorphous fluoropolymer having a refractive index lessthan the refractive index of water and having an absorption coefficientof a magnitude such that when said conduit is filled with water, visiblelight and ultra-violet light can be transmitted, substantially withoutloss, along an axis of said conduit by attenuated total reflection andvisible light and ultra-violet light are substantially completelyabsorbed in passage through the wall of said conduit.
 24. A method ofperforming photometric analysis of a liquid sample comprising:introducing said liquid sample into a conduit having a wall formed of anamorphous fluoropolymer having a refractive index less than a refractiveindex of water and having an absorption coefficient sufficient tosubstantially attenuate light propagation through said wall; shininglight axially onto said conduit filled with sample liquid; receivinglight transferred through said liquid sample at a detector; anddetermining a concentration of said sample in said liquid by lightabsorption by the sample, whereby linearity of detection is improved.25. The method of claim 24 whereby said light shining axially is at anexcitation wavelength and said light transferred is emitted fluorescenceor Raman scattered light used to determine a sample concentration and asample identity, such that unwanted scattered excitation light enteringsaid conduit wall is suppressed and does not reach a detector.
 26. A setof light-guiding flowcells for a fluid comprising: a set of channelsformed in a substrate of material having an index of refraction lowerthan an index of said fluid and an absorption coefficient sufficient tosubstantially attenuate light propagating through said material; a setof channel covers formed in a section of material having an index ofrefraction identical to said substrate material and an absorptioncoefficient sufficient to substantially attenuate light propagatingthrough said material, said set of channel covers fixed to said set ofchannels forming a set of covered channels; at least one covered channelhaving a fluid inlet and outlet port configured off an axis of saidcovered channel; at least one covered channel having a source of lightat a light entrance end and having a light exit end; whereby said atleast one covered channel and fluid effect guiding of said light byattenuated total reflection and said at least one covered channel hasminimal absorptive effect on said light guided by internal reflection.27. The set of light-guiding flowcells of claim 26 further comprising anoptical fiber bonded to at least one covered channel providing saidlight source.
 28. The set of light-guiding flowcells of claim 26 wherebythe material forming said closed channels prevents light crosstalkbetween flowcells.
 28. The set of light-guiding flowcells of claim 26wherein at least one covered channel is configured as a separationcolumn.
 29. The set of light-guiding flowcells of claim 26 wherein atleast one covered channel is configured as a fluidic connection.
 30. Theset of light-guiding flowcells of claim 25 wherein said substrate andsaid section are formed of Teflon® AF fluoropolymer doped with a blackdopant at a concentration of between 0.01-0.1% of the fluoropolymer.